Cancer is an uncontrolled growth of abnormal cells. It is found in animals as well as humans. There are many different types of cancer that are found in animals, symptoms are often similar to those in people (eg. abnormal swelling, unexplained weight loss, lethargy / reluctance to exercise etc). Cancer is one of the leading causes of death in companion animals such as dogs and cats, it is particularly common in animals that live 10 years or longer. If treatment is appropriate this may include chemotherapy and surgery or radiotherapy.
Zamani-Ahmadmahmudi M, Aghasharif S, Ilbeigi K Prognostic efficacy of the human B-cell lymphoma prognostic genes in predicting disease-free survival (DFS) in the canine counterpart. BMC Vet Res. 2017; 13(1):17 [PubMed] Free Access to Full ArticleRelated Publications
BACKGROUND: Canine B-cell lymphoma is deemed an ideal model of human non-Hodgkin's lymphoma where the lymphomas of both species share similar clinical features and biological behaviors. However there are some differences between tumor features in both species. In the current study, we sought to evaluate the prognostic efficacy of human B-cell lymphoma prognostic gene signatures in canine B-cell lymphoma. METHODS: The corresponding probe sets of 36 human B-cell lymphoma prognostic genes were retrieved from 2 canine B-cell lymphoma microarray datasets (GSE43664 and GSE39365) (76 samples), and prognostic probe sets were thereafter detected using the univariate and multivariate Cox proportional-hazard model and the Kaplan-Meier analysis. The two datasets were employed both as training sets and as external validation sets for each other. Results were confirmed using quantitative real-time PCR (qRT-PCR) analysis. RESULTS: In the univariate analysis, CCND1, CCND2, PAX5, CR2, LMO2, HLA-DQA1, P53, CD38, MYC-N, MYBL1, and BIRCS5 were associated with longer disease-free survival (DFS), while CD44, PLAU, and FN1 were allied to shorter DFS. However, the multivariate Cox proportional-hazard analysis confirmed CCND1 and BIRCS5 as prognostic genes for canine B-cell lymphoma. qRT-PCR used for verification of results indicated that expression level of CCND1 was significantly higher in B-cell lymphoma patients with the long DFS than ones with the short DFS, while expression level of BIRCS5 wasn't significantly different between two groups. CONCLUSION: Our results confirmed CCND1 as important gene that can be used as a potential predictor in this tumor type.
Infiltrating cells of the immune system are widely accepted to be generic constituents of tumor microenvironment. It has been well established that the development of mammary cancer, both in humans and in dogs, is associated with alterations in numbers and functions of immune cells at the sites of tumor progression. These tumor infiltrating immune cells seem to exhibit exclusive phenotypic and functional characteristics and mammary cancer cells can take advantage of signaling molecules released by them. Cancer related inflammation has an important role in mammary carcinogenesis, contributing to the acquisition of core hallmark capabilities that allow cancer cells to survive, proliferate, and disseminate. Indeed, recent studies in human breast cancer and in canine mammary tumors have identified a growing list of signaling molecules released by inflammatory cells that serve as effectors of their tumor-promoting actions. These include the COX-2, the tumor EGF, the angiogenic VEGF, other proangiogenic factors, and a large variety of chemokines and cytokines that amplify the inflammatory state. This review describes the intertwined signaling pathways shared by T-lymphocytic/macrophage infiltrates and important tissue biomarkers in both human and dog mammary carcinogenesis.
Cancer stem cells (CSC) represent a small subpopulation of cells in malignant tumors that possess the unique ability to self-renew, differentiate and resist chemo- and radiotherapy. These cells have been postulated to be the basis for some of the difficulties in treating cancer, and therefore, numerous approaches have been developed to specifically target and eliminate CSC in diverse types of cancer, including breast cancer. Spontaneously occurring mammary tumors in canines share clinical and molecular similarities with the human counterpart, making the dog a potentially powerful model for the study of human breast cancer and clinical trials. Studies focused on canine mammary CSC might therefore enhance our understanding of the biology and possible treatment of the disease in both dogs and humans. In this review, we discuss various approaches currently in use to isolate and characterize canine mammary CSC.
Gupta SK, Yadav PK, Tiwari AK, et al. Poly (I:C) enhances the anti-tumor activity of canine parvovirus NS1 protein by inducing a potent anti-tumor immune response. Tumour Biol. 2016; 37(9):12089-12102 [PubMed] Related Publications
The canine parvovirus NS1 (CPV2.NS1) protein selectively induces apoptosis in the malignant cells. However, for an effective in vivo tumor treatment strategy, an oncolytic agent also needs to induce a potent anti-tumor immune response. In the present study, we used poly (I:C), a TLR3 ligand, as an adjuvant along with CPV2.NS1 to find out if the combination can enhance the oncolytic activity by inducing a potent anti-tumor immune response. The 4T1 mammary carcinoma cells were used to induce mammary tumor in Balb/c mice. The results suggested that poly (I:C), when given along with CPV2.NS1, not only significantly reduced the tumor growth but also augmented the immune response against tumor antigen(s) as indicated by the increase in blood CD4+ and CD8+ counts and infiltration of immune cells in the tumor tissue. Further, blood serum analysis of the cytokines revealed that Th1 cytokines (IFN-γ and IL-2) were significantly upregulated in the treatment group indicating activation of cell-mediated immune response. The present study reports the efficacy of CPV2.NS1 along with poly (I:C) not only in inhibiting the mammary tumor growth but also in generating an active anti-tumor immune response without any visible toxicity. The results of our study may help in developing CPV2.NS1 and poly (I: C) combination as a cancer therapeutic regime to treat various malignancies.
BACKGROUND: Current studies report that aberrations in epigenetic regulators or chromatin modifications are related to tumor development and maintenance. EZH2 (Enhancer of zeste homolog 2) is one of the catalytic subunits of Polycomb repressive complex 2, a crucial epigenetic regulator. EZH2 has a master regulatory function in such processes as cell proliferation, stem cell differentiation, and early embryogenesis. In humans, EZH2 is linked to oncogenic function in several carcinomas, including breast cancer, and dysregulation of EZH2 has been particularly associated with loss of differentiation and the development of poorly differentiated breast cancer. In our present study, we were interested in determining whether EZH2 is increased in canine mammary tumors, which show similarities to human breast cancer. RESULTS: Investigation of the expression of EZH2 in canine mammary tumors revealed that EZH2 protein was overexpressed in canine mammary carcinomas, as in human breast cancer. In addition, the immunohistochemical expression level of EZH2 was associated with the degree of malignancy in canine mammary carcinoma. This is the first report to describe EZH2 expression in canine mammary tumors. CONCLUSIONS: Because the expression of EZH2 was similar in canine mammary carcinoma and human breast cancer, spontaneous canine mammary tumors may be a suitable model for studying EZH2 and treatment development.
Sirivisoot S, Techangamsuwan S, Tangkawattana S, Rungsipipat A Application of Immunophenotyping and Heteroduplex Polymerase Chain Reaction (hPARR) for Diagnosis of Canine Lymphomas. Asian Pac J Cancer Prev. 2016; 17(6):2909-16 [PubMed] Related Publications
BACKGROUND: Canine malignant lymphoma is classified into B- or T-cell origin, as in the human case. Due to differences in prognosis, a suitable method needs to be developed for lineage identification. AIMS: To determine the accuracy of immunophenotypic and molecular information between three methods: immunocytochemistry (ICC), immunohistochemistry (IHC) and heteroduplex polymerase chain reaction for antigen receptor rearrangements (hPARR) in spontaneous canine lymphomas. MATERIALS AND METHODS: Peripheral blood, fine needle aspiration and tissue biopsies from enlarged peripheral lymph nodes prior to treatment of 28 multicentric lymphoma patients were collected. Cytopathology and histopathology were examined and classified using the updated Kiel and WHO classifications, respectively. Anti-Pax5 and anti-CD3 antibodies as B- and T-cell markers were applied for immunophenotyping by ICC and IHC. Neoplastic lymphocytes from lymph node and white blood cell pellets from peripheral blood were evaluated by hPARR. RESULTS: In this study, low grade B-cell lymphoma accounted for 25% (7/28), high grade B-cell lymphoma for 64.3% (18/28) and high grade T-cell lymphoma for 10.7% (3/28). According to the WHO classification, 50% of all cases were classified as diffuse large B-cell lymphoma. In addition, ICC showed concordant results with IHC; all B-cell lymphomas showed Pax5+/CD3, and all T-cell lymphomas exhibited Pax5-/CD3+. In contrast to hPARR, 12 B-cell lymphomas featured the IgH gene; seven presented the TCRγ gene; five cases showed both IgH and TCRγ genes, and one case were indeterminate. Three T-cell lymphomas showed the TCRγ gene. The percentage agreement between hPARR and ICC/IHC was 60%. CONCLUSIONS: Immunophenotyping should not rely on a single method. ICC or IHC with hPARR should be used concurrently for immunophenotypic diagnosis in canine lymphomas.
Canine inflammatory mammary cancer (IMC) shares clinical and histopathological characteristics with human inflammatory breast cancer (IBC) and has been proposed as a good model for studying the human disease. The aim of this study was to evaluate the capacity of female and male mice to reproduce IMC and IBC tumors and identify the hormonal tumor environment. To perform the study sixty 6-8-week-old male and female mice were inoculated subcutaneously with a suspension of 10(6)IPC-366 and SUM149 cells. Tumors and serum were collected and used for hormonal analysis. Results revealed that IPC-366 reproduced tumors in 90% of males inoculated after 2 weeks compared with 100% of females that reproduced tumor at the same time. SUM149 reproduced tumors in 40% of males instead of 80% of females that reproduced tumors after 4 weeks. Both cell lines produce distant metastasis in lungs being higher than the metastatic rates in females. EIA analysis revealed that male tumors had higher T and SO4E1 concentrations compared to female tumors. Serum steroid levels were lower than those found in tumors. In conclusion, IBC and IMC male mouse model is useful as a tool for IBC research and those circulating estrogens and intratumoral hormonal levels are crucial in the development and progression of tumors.
MacDiarmid JA, Langova V, Bailey D, et al. Targeted Doxorubicin Delivery to Brain Tumors via Minicells: Proof of Principle Using Dogs with Spontaneously Occurring Tumors as a Model. PLoS One. 2016; 11(4):e0151832 [PubMed] Free Access to Full ArticleRelated Publications
BACKGROUND: Cytotoxic chemotherapy can be very effective for the treatment of cancer but toxicity on normal tissues often limits patient tolerance and often causes long-term adverse effects. The objective of this study was to assist in the preclinical development of using modified, non-living bacterially-derived minicells to deliver the potent chemotherapeutic doxorubicin via epidermal growth factor receptor (EGFR) targeting. Specifically, this study sought to evaluate the safety and efficacy of EGFR targeted, doxorubicin loaded minicells (designated EGFRminicellsDox) to deliver doxorubicin to spontaneous brain tumors in 17 companion dogs; a comparative oncology model of human brain cancers. METHODOLOGY/PRINCIPLE FINDINGS: EGFRminicellsDox were administered weekly via intravenous injection to 17 dogs with late-stage brain cancers. Biodistribution was assessed using single-photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI). Anti-tumor response was determined using MRI, and blood samples were subject to toxicology (hematology, biochemistry) and inflammatory marker analysis. Targeted, doxorubicin-loaded minicells rapidly localized to the core of brain tumors. Complete resolution or marked tumor regression (>90% reduction in tumor volume) were observed in 23.53% of the cohort, with lasting anti-tumor responses characterized by remission in three dogs for more than two years. The median overall survival was 264 days (range 49 to 973). No adverse clinical, hematological or biochemical effects were observed with repeated administration of EGFRminicellsDox (30 to 98 doses administered in 10 of the 17 dogs). CONCLUSIONS/SIGNIFICANCE: Targeted minicells loaded with doxorubicin were safely administered to dogs with late stage brain cancer and clinical activity was observed. These findings demonstrate the strong potential for clinical applications of targeted, doxorubicin-loaded minicells for the effective treatment of patients with brain cancer. On this basis, we have designed a Phase 1 clinical study of EGFR-targeted, doxorubicin-loaded minicells for effective treatment of human patients with recurrent glioblastoma.
Vascellari M, Capello K, Carminato A, et al. Incidence of mammary tumors in the canine population living in the Veneto region (Northeastern Italy): Risk factors and similarities to human breast cancer. Prev Vet Med. 2016; 126:183-9 [PubMed] Related Publications
Although mammary gland tumors (MT) are the most-common type of tumor in intact female dogs, there is little information about their incidence in dog population. Data on MT in female dogs was retrieved from the Animal Tumor registry of dogs and cats of Venice and Vicenza provinces during 2005-2013 and was analyzed to visualize crude incidence rates by breed and across age categories. Overall, 2744 mammary tumors were reported accounting for 54% of all tumors in female dogs. The annual incidence rate (IR) was 250 cases per 100,000 dogs. The most frequent malignant tumors were complex carcinomas, consisting of both epithelial and myoepithelial tissues (IR=71.89), and simple carcinomas (IR=62.59). The MT incidence rate increased through the study period; particularly in the last 4 years, and malignant neoplasms occurred more frequently (70%) than the benign counterparts (30%). Seventy-four percent of tumors were diagnosed in intact females, and the mean age at diagnosis was significantly higher for spayed dogs than for intact ones. MT were less frequent in dogs younger than 6 years and increased up to approximately 60% for ages between 8 and 13 years. The purebred dogs had a higher probability to have a malignant neoplasm than mixed-breed dogs, particularly in dogs younger than 7 years, and the Samoyed, Dobermann, Schnauzer and Yorkshire Terrier breeds were more inclined to develop malignant MT. The incidence of MT in dogs is increasing, and IRs are comparable to that in women. The epidemiological similarities between dogs and women support the validity of canine MT as a model for human breast cancer.
Gonçalves Ndo N, Colombo J, Lopes JR, et al. Effect of Melatonin in Epithelial Mesenchymal Transition Markers and Invasive Properties of Breast Cancer Stem Cells of Canine and Human Cell Lines. PLoS One. 2016; 11(3):e0150407 [PubMed] Free Access to Full ArticleRelated Publications
Cancer stem cells (CSCs) have been associated with metastasis and therapeutic resistance and can be generated via epithelial mesenchymal transition (EMT). Some studies suggest that the hormone melatonin acts in CSCs and may participate in the inhibition of the EMT. The objectives of this study were to evaluate the formation of mammospheres from the canine and human breast cancer cell lines, CMT-U229 and MCF-7, and the effects of melatonin treatment on the modulation of stem cell and EMT molecular markers: OCT4, E-cadherin, N-cadherin and vimentin, as well as on cell viability and invasiveness of the cells from mammospheres. The CMT-U229 and MCF-7 cell lines were subjected to three-dimensional culture in special medium for stem cells. The phenotype of mammospheres was first evaluated by flow cytometry (CD44(+)/CD24(low/-) marking). Cell viability was measured by MTT colorimetric assay and the expression of the proteins OCT4, E-cadherin, N-cadherin and vimentin was evaluated by immunofluorescence and quantified by optical densitometry. The analysis of cell migration and invasion was performed in Boyden Chamber. Flow cytometry proved the stem cell phenotype with CD44(+)/CD24(low/-) positive marking for both cell lines. Cell viability of CMT-U229 and MCF-7 cells was reduced after treatment with 1mM melatonin for 24 h (P<0.05). Immunofluorescence staining showed increased E-cadherin expression (P<0.05) and decreased expression of OCT4, N-cadherin and vimentin (P<0.05) in both cell lines after treatment with 1 mM melatonin for 24 hours. Moreover, treatment with melatonin was able to reduce cell migration and invasion in both cell lines when compared to control group (P<0.05). Our results demonstrate that melatonin shows an inhibitory role in the viability and invasiveness of breast cancer mammospheres as well as in modulating the expression of proteins related to EMT in breast CSCs, suggesting its potential anti-metastatic role in canine and human breast cancer cell lines.
HER2 is overexpressed in about 30% of feline mammary carcinomas (FMC) and in 15-30% of breast cancers. Women with HER2-positive breast tumors are associated with shorter survival. This study aimed to optimize the detection and quantification of serum HER2 (sHER2) in cats and to evaluate its potential in diagnosing cats with mammary carcinomas (MC) overexpressing HER2. A prospective study was conducted in 60 queens showing MC and 20 healthy animals. Pre-operative serum samples were collected for sHER2 quantification using two immunoassays: ELISA and Dot blot assay. sHER2 levels were compared with tissue HER2 status assessed by immunohistochemistry. Queens with FMC showed significantly higher mean levels of sHER2 by both ELISA and Dot blot assay. A significant difference in the sHER2 levels was also found between cats with HER2-positive MC and those with low-expressing HER2 MC. A significant correlation between sHER2 levels and tumor HER2 status was also found, particularly when ELISA was used (r = 0.58, p < 0.0001). The value of 10 ng/ml was proposed as the optimal cutoff for both immunoassays by ROC analysis. Like in humans, sHER2 levels are increased in cats with MC HER2-positive, strongly suggesting that evaluation of sHER2 levels can be very useful in feline oncology. The results show that ELISA and Dot blot assay can replace the immunohistochemistry technique, due to their efficacy and lower costs for diagnostic purposes and for monitoring the response to anti-HER2 therapies in cats.
Hood G, Laufer-Amorim R, Fonseca-Alves CE, Palmieri C Overexpression of Ephrin A3 Receptor in Canine Prostatic Carcinoma. J Comp Pathol. 2016 Feb-Apr; 154(2-3):180-5 [PubMed] Related Publications
Ephrin A3 (EphA3), a member of the ephrin receptor tyrosine kinase family, is involved in a variety of functions in normal cells, especially during embryonic development, and alterations in its expression profile have been observed in several human cancers. However, there are no reports of the expression of EphA3 in normal, hyperplastic or neoplastic canine prostate tissue or in other types of canine tumours. Six normal, 15 hyperplastic and 21 neoplastic canine prostates were examined immunohistochemically with a polyclonal antibody specific for human EphA3. The percentage of positive cells in all prostatic carcinomas was increased, with a mean of 89.28 ± 5.18% compared with normal (9.17 ± 6.72%) and hyperplastic prostates (20.00 ± 8.28%). EphA3 expression was not correlated with the histological subtypes of prostate cancer or with the Gleason score. The increase in EphA3 expression in canine prostatic carcinomas suggests the involvement of this receptor in prostatic carcinogenesis and its potential use as a target for new therapeutic strategies.
BACKGROUND: Genomics-based predictors of drug response have the potential to improve outcomes associated with cancer therapy. Osteosarcoma (OS), the most common primary bone cancer in dogs, is commonly treated with adjuvant doxorubicin or carboplatin following amputation of the affected limb. We evaluated the use of gene-expression based models built in an intra- or interspecies manner to predict chemosensitivity and treatment outcome in canine OS. Models were built and evaluated using microarray gene expression and drug sensitivity data from human and canine cancer cell lines, and canine OS tumor datasets. The "COXEN" method was utilized to filter gene signatures between human and dog datasets based on strong co-expression patterns. Models were built using linear discriminant analysis via the misclassification penalized posterior algorithm. RESULTS: The best doxorubicin model involved genes identified in human lines that were co-expressed and trained on canine OS tumor data, which accurately predicted clinical outcome in 73 % of dogs (p = 0.0262, binomial). The best carboplatin model utilized canine lines for gene identification and model training, with canine OS tumor data for co-expression. Dogs whose treatment matched our predictions had significantly better clinical outcomes than those that didn't (p = 0.0006, Log Rank), and this predictor significantly associated with longer disease free intervals in a Cox multivariate analysis (hazard ratio = 0.3102, p = 0.0124). CONCLUSIONS: Our data show that intra- and interspecies gene expression models can successfully predict response in canine OS, which may improve outcome in dogs and serve as pre-clinical validation for similar methods in human cancer research.
Atherton MJ, Morris JS, McDermott MR, Lichty BD Cancer immunology and canine malignant melanoma: A comparative review. Vet Immunol Immunopathol. 2016; 169:15-26 [PubMed] Related Publications
Oral canine malignant melanoma (CMM) is a spontaneously occurring aggressive tumour with relatively few medical treatment options, which provides a suitable model for the disease in humans. Historically, multiple immunotherapeutic strategies aimed at provoking both innate and adaptive anti-tumour immune responses have been published with varying levels of activity against CMM. Recently, a plasmid DNA vaccine expressing human tyrosinase has been licensed for the adjunct treatment of oral CMM. This article reviews the immunological similarities between CMM and the human counterpart; mechanisms by which tumours evade the immune system; reasons why melanoma is an attractive target for immunotherapy; the premise of whole cell, dendritic cell (DC), viral and DNA vaccination strategies alongside preliminary clinical results in dogs. Current "gold standard" treatments for advanced human malignant melanoma are evolving quickly with remarkable results being achieved following the introduction of immune checkpoint blockade and adoptively transferred cell therapies. The rapidly expanding field of cancer immunology and immunotherapeutics means that rational targeting of this disease in both species should enhance treatment outcomes in veterinary and human clinics.
Herranz C, Fernández F, Martín-Ibáñez R, et al. Spontaneously Arising Canine Glioma as a Potential Model for Human Glioma. J Comp Pathol. 2016 Feb-Apr; 154(2-3):169-79 [PubMed] Related Publications
Human gliomas are malignant brain tumours that carry a poor prognosis and are composed of a heterogeneous population of cells. There is a paucity of animal models available for study of these tumours and most have been created by genetic modification. Spontaneously arising canine gliomas may provide a model for the characterization of the human tumours. The present study shows that canine gliomas form a range of immunohistochemical patterns that are similar to those described for human gliomas. The in-vitro sphere assay was used to analyze the expansion and differentiation potential of glioma cells taken from the periphery and centre of canine tumours. Samples from the subventricular zone (SVZ) and contralateral parenchyma were used as positive and negative controls, respectively. The expansion potential for all of these samples was low and cells from only three cultures were expanded for six passages. These three cultures were derived from high-grade gliomas and the cells had been cryopreserved. Most of the cells obtained from the centre of the tumours formed spheres and were expanded, in contrast to samples taken from the periphery of the tumours. Spheres were also formed and expanded from two areas of apparently unaffected brain parenchyma. The neurogenic SVZ contralateral samples also contained progenitor proliferating cells, since all of them were expanded for three to five passages. Differentiation analysis showed that all cultured spheres were multipotential and able to differentiate towards both neurons and glial cells. Spontaneously arising canine gliomas might therefore constitute an animal model for further characterization of these tumours.
Morais CS, Affonso PR, Bitencourt JA, Wenceslau AA Direct preparation protocol to obtain mitotic chromosomes from canine mammary tumors. Genet Mol Res. 2015; 14(4):19389-95 [PubMed] Related Publications
Currently, mammary neoplasms in female canines are a serious problem in veterinary clinics. In addition, the canine species is an excellent disease model for human oncology because of the biological and genetic similarities between the species. Cytogenetics has allowed further study of the characterization of neoplasms in canines. We hypothesized that the use of a direct preparation protocol for mitotic chromosome analysis would provide a simple and low cost protocol for use in all laboratories. The objective of this method is to display in a few hours of dividing cells just like the time of collection since cell division in tissue can be obtained. Ten female canines with the spontaneous occurrence of mammary neoplasia were used to test a pioneering direct preparation protocol to obtain mitotic chromosomes. The excised breast tumor tissue fragments were subjected to the protocol consisting of treatment with colchicine, treatment with hypotonic solution, and fixation. Mitotic chromosomes were absent in cell suspensions of only two samples among the 10 materials analyzed, based on the analysis of five blades for each preparation obtained. So, the cell suspension obtained allowed for the observation of eight tissue samples viable for cytogenetic analysis, five of which had excellent numbers of mitotic chromosomes. However, the technique was unsuccessful in producing high-quality cell suspensions because of inadequate condensation and scattering of chromosomes. While adjustments to methodological procedures are needed, this protocol represents a low cost and simplified method to study the cytogenetics of canine tumors.
The oncolytic effects of reovirus in various cancers have been proven in many clinical trials in human medicine. Oncolytic virotherapy using reovirus for canine cancers is being developed in our laboratory. The objective of this study was to examine the synergistic anti-cancer effects of a combination of reovirus and low doses of various chemotherapeutic agents on mammary gland tumors (MGTs) in dogs. The first part of this study demonstrated the efficacy of reovirus in canine MGTs in vitro and in vivo. Reovirus alone exerted significant cell death by means of caspase-dependent apoptosis in canine MGT cell lines. A single injection of reovirus impeded growth of canine MGT tumors in xenografted mice, but was insufficient to induce complete tumor regression. The second part of this study highlighted the anti-tumor effects of reovirus in combination with low doses of paclitaxel, carboplatin, gemcitabine, or toceranib. Enhanced synergistic activity was observed in the MGT cell line treated concomitantly with reovirus and in all the chemotherapeutic agents except toceranib. In addition, combining reovirus with paclitaxel or gemcitabine at half dosage of half maximal inhibitory concentration (IC50) enhanced cytotoxicity by activating caspase 3. Our data suggest that the combination of reovirus and low dose chemotherapeutic agents provides an attractive option in canine cancer therapy.
Research on oncolytic viruses has mostly been directed towards the treatment of solid tumors, which has yielded limited information regarding their activity in hematological cancer. It has also been directed towards the treatment of humans, yet veterinary medicine may also benefit. Several strains of the Newcastle disease virus (NDV) have been used as oncolytics in vitro and in a number of in vivo experiments. We studied the cytolytic effect of NDV-MLS, a low virulence attenuated lentogenic strain, on a human large B-cell lymphoma cell line (SU-DHL-4), as well as on primary canine-derived B-cell lymphoma cells, and compared them to healthy peripheral blood mononuclear cells (PBMC) from both humans and dogs. NDV-MLS reduced cell survival in both human (42% ± 5%) and dog (34% ± 12%) lymphoma cells as compared to untreated controls. No significant effect on PBMC was seen. Cell death involved apoptosis as documented by flow-cytometry. NDV-MLS infections of malignant lymphoma tumors in vivo in dogs were confirmed by electron microscopy. Early (24 h) biodistribution of intravenous injection of 1 × 10(12) TCID50 (tissue culture infective dose) in a dog with T-cell lymphoma showed viral localization only in the kidney, the salivary gland, the lung and the stomach by immunohistochemistry and/or endpoint PCR. We conclude that NDV-MLS may be a promising agent for the treatment of lymphomas. Future research is needed to elucidate the optimal therapeutic regimen and establish appropriate biosafety measures.
Sánchez-Céspedes R, Millán Y, Guil-Luna S, et al. Myoepithelial cells in canine mammary tumours. Vet J. 2016; 207:45-52 [PubMed] Related Publications
Mammary tumours are the most common neoplasms of female dogs. Compared to mammary tumours of humans and cats, myoepithelial (ME) cell involvement is common in canine mammary tumours (CMT) of any subtype. Since ME cell involvement in CMT influences both histogenetic tumour classification and prognosis, correct identification of ME cells is important. This review describes immunohistochemical methods for identification of canine mammary ME cells used in vivo. In addition, phenotypic and genotypic methods to isolate ME cells for in vitro studies to analyse tumour-suppressor protein production and gene expression are discussed. The contribution of ME cells to both histogenetic classifications and the prognosis of CMT is compared with other species and the potential use of ME cells as a method to identify carcinoma in situ is discussed.
Takanosu M, Amano S, Kagawa Y Analysis of c-KIT exon 11 mutations in canine gastrointestinal stromal tumours. Vet J. 2016; 207:118-23 [PubMed] Related Publications
The aim of this study was to determine the type and frequency of c-KIT exon 11 mutations in canine gastrointestinal stromal tumours (GISTs) and investigate the association between the c-KIT mutation status and KIT immunohistochemical staining pattern. Mutations in exon 11 of c-KIT were examined in 46 formalin-fixed paraffin-embedded canine GISTs using PCR of genomic DNA and reverse transcription-PCR (RT-PCR) of cDNA. Exon 11 c-KIT mutations were detected in 15/46 (32.6%) cases by conventional PCR and 34/46 (73.9%) cases by RT-PCR; the mutation detection rate was significantly higher for RT-PCR (P = 0.004, Fisher's exact test). Ten different mutations, including deletion, internal tandem duplication and point mutations, were identified by RT-PCR. Immunohistochemistry was performed using an anti-KIT antibody; diffuse KIT staining was detected in the tumour cell cytoplasm in 32/46 (69.6%) cases and partial or stippled cytoplasmic staining of KIT was observed in 14/46 (30.4%) cases. Neither pattern was significantly associated with c-KIT exon 11 mutation status (P = 1.000, chi-square test). These data indicate that c-KIT exon 11 mutations occur frequently in canine GISTs, similar to human GISTs; however, there is no association between c-KIT mutations and the KIT expression pattern in canine GISTs. This study suggests that RT-PCR is more sensitive than conventional PCR for the detection of c-KIT mutations in canine GISTs.
Beffagna G, Sacchetto R, Cavicchioli L, et al. A preliminary investigation of the role of the transcription co-activators YAP/TAZ of the Hippo signalling pathway in canine and feline mammary tumours. Vet J. 2016; 207:105-11 [PubMed] Related Publications
Breast cancer is the most common cancer in women worldwide. Cancer metastases are responsible for the high mortality rate. A small but distinct subset of cells, cancer stem cells (CSCs), have the capacity to self-renew, initiate tumour formation, and develop metastases. The CSC content in human breast cancer correlates with the Hippo tumour suppressor signalling pathway. Specifically, the activity of YAP/TAZ, transcription co-activators of the Hippo pathway, sustains the self-renewal and tumour-initiation capacities of CSCs. Little is known about YAP/TAZ in canine and feline mammary tumours, which are very common tumours. The preliminary aim of the study was to investigate the expression of YAP/TAZ in canine and feline mammary tumours by Western blot and immunohistochemistry. Increased cytoplasmic and nuclear expression of YAP/TAZ was observed in all carcinomas compared to normal tissues, indicating neoplastic deregulation of the Hippo pathway. Nuclear expression significantly increased in grade III (high grade carcinomas) compared to grade I (low grade carcinomas) tumours, suggesting that YAP/TAZ play a role in the increased aggressiveness of these tumours. Moreover, different scoring systems for immunohistochemical analyses were compared and the H index and the Allred scores were the most significant. In conclusion, YAP/TAZ are expressed in aggressive canine and feline mammary tumours as reported in some human cancers. Further studies might better elucidate the role of the Hippo pathway in prognosis and as a target for new therapies. In addition, tumours in dogs and cats may be a useful model to study this pathway.
Yoel U, Gopas J, Ozer J, et al. Canine Scent Detection of Volatile Elements, Characteristic of Malignant Cells, in Cell Cultures. Isr Med Assoc J. 2015; 17(9):567-70 [PubMed] Related Publications
BACKGROUND: In recent years several reports have been published describing dogs' ability to detect, by scent, patients with cancer. This ability is based on the sniffing of volatile organic elements that are secreted by malignant cells or react to them. OBJECTIVES: To evaluate the ability of trained dogs to detect breast cancer cell cultures (MCF7) compared to the control pseudo-normal keratinocyte cell line (HaCaT), and to detect melanoma (BG) and type 2 epithelial lung carcinoma (A549) malignant cell cultures to which they were not previously exposed in the course of their training. METHODS: Cell cultures were prepared in a standard manner. Two Belgian Shepherd dogs were trained and then tested in a single-blind test (for dogs and trainers) on their ability to detect the "target specimen," a MCF7 breast cancer cell culture. Following this, the ability of the dogs to detect cancer cell cultures that they were not previously exposed to (i.e., A549, BG) was tested. In each test round, four specimens placed in identical blocks were arranged in a line with one meter between them: one target specimen (MCF7, A549, BG), two control specimens (HaCaT), and a sample containing cell culture medium only. RESULTS: The two dogs picked out all the target specimens of MCF7 breast cancer cell cultures that they were trained to detect (10/10) as well as all the target specimens that they were not previously exposed to [A549 (5/5) and BG (5/5)], but did not pick out the control specimens or the cell culture medium. Thus, the sensitivity, specificity, and positive and negative predictive values for both dogs were 100%. CONCLUSIONS: The results of this study support the assumption that cancer cells have a unique odor pattern, and that this odor pattern is common to different types of cancer.
Spontaneous cancers in client-owned dogs closely recapitulate their human counterparts with respect to clinical presentation, histological features, molecular profiles, and response and resistance to therapy, as well as the evolution of drug-resistant metastases. In several instances the incorporation of dogs with cancer into the preclinical development path of cancer therapeutics has influenced outcome by helping to establish pharmacokinetic/pharmacodynamics relationships, dose/regimen, expected clinical toxicities, and ultimately the potential for biologic activity. As our understanding regarding the molecular drivers of canine cancers has improved, unique opportunities have emerged to leverage this spontaneous model to better guide cancer drug development so that therapies likely to fail are eliminated earlier and therapies with true potential are optimized prior to human studies. Both pets and people benefit from this approach, as it provides dogs with access to cutting-edge cancer treatments and helps to insure that people are given treatments more likely to succeed.
Zorzan E, Hanssens K, Giantin M, et al. Mutational Hotspot of TET2, IDH1, IDH2, SRSF2, SF3B1, KRAS, and NRAS from Human Systemic Mastocytosis Are Not Conserved in Canine Mast Cell Tumors. PLoS One. 2015; 10(11):e0142450 [PubMed] Free Access to Full ArticleRelated Publications
INTRODUCTION: Both canine cutaneous mast cell tumor (MCT) and human systemic mastocytosis (SM) are characterized by abnormal proliferation and accumulation of mast cells in tissues and, frequently, by the presence of activating mutations in the receptor tyrosine kinase V-Kit Hardy-Zuckerman 4 Feline Sarcoma Viral Oncogene Homolog (c-KIT), albeit at different incidence (>80% in SM and 10-30% in MCT). In the last few years, it has been discovered that additional mutations in other genes belonging to the methylation system, the splicing machinery and cell signaling, contribute, with c-KIT, to SM pathogenesis and/or phenotype. In the present study, the mutational profile of the Tet methylcytosine dioxygenase 2 (TET2), the isocitrate dehydrogenases 1 and 2 (IDH1 and IDH2), the serine/arginine-rich splicing factor 2 (SRSF2), the splicing factor 3b subunit 1 (SF3B1), the Kirsten rat sarcoma viral oncogene homolog (KRAS) and the neuroblastoma RAS viral oncogene homolog (NRAS), commonly mutated in human myeloid malignancies and mastocytosis, was investigated in canine MCTs. METHODS: Using the Sanger sequencing method, a cohort of 75 DNA samples extracted from MCT biopsies already investigated for c-KIT mutations were screened for the "human-like" hot spot mutations of listed genes. RESULTS: No mutations were ever identified except for TET2 even if with low frequency (2.7%). In contrast to what is observed in human TET2 no frame-shift mutations were found in MCT samples. CONCLUSION: Results obtained in this preliminary study are suggestive of a substantial difference between human SM and canine MCT if we consider some target genes known to be involved in the pathogenesis of human SM.
Sox9 is a master regulatory gene involved in developmental processes, stem cells maintenance and tumorigenesis. This gene is expressed in healthy skin but even in several skin neoplasms, where its expression patterns often resembles those of the developing hair follicle. In this study, samples from eleven different types of canine skin neoplasms (squamous papilloma, squamous cell carcinoma, infundibular keratinizing acanthoma, inferior tricholemmoma, isthmic tricholemmoma, trichoblastoma, trichoepitelioma, malignant trichoepitelioma, pilomatricoma, subungual keratoacanthoma, subungual squamous cell carcinoma) were immunohistochemically stained and evaluated for Sox9 with the aim to correlate tumor phenotype with molecular characteristics that may help to better define tumor development, contribute to its diagnosis and clinical management. Keratoacanthoma excluded, all the skin neoplasms examined showed a variable positivity to Sox9, especially in the basal layers, but with major intensity in neoplasms developing from the bulge region of the hair follicle, as trichoblastoma. According to our results, Sox9 could be employed as a stem cell marker to better assess the role of stem cells in canine epidermal and follicular tumors.
Ebisawa M, Iwano H, Nishikawa M, et al. Significance of caveolin-1 and matrix metalloproteinase 14 gene expression in canine mammary tumours. Vet J. 2015; 206(2):191-6 [PubMed] Related Publications
Canine mammary tumours (CMTs) are the most common neoplasms affecting female dogs. There is an urgent need for molecular biomarkers that can detect early stages of the disease in order to improve accuracy of CMT diagnosis. The aim of this study was to examine whether caveolin-1 (Cav-1) and matrix metalloproteinase 14 (MMP14) are associated with CMT histological malignancy and invasion. Sixty-five benign and malignant CMT samples and six normal canine mammary glands were analysed using quantitative reverse transcription-polymerase chain reaction. Cav-1 and MMP14 genes were highly expressed in CMT tissues compared to normal tissues. Cav-1 especially was overexpressed in malignant and invasive CMT tissues. When a CMT cell line was cultured on fluorescent gelatin-coated coverslips, localisation of Cav-1 was observed at invadopodia-mediated degradation sites of the gelatin matrix. These findings suggest that Cav-1 may be involved in CMT invasion and that the markers may be useful for estimating CMT malignancy.
Dhawan D, Paoloni M, Shukradas S, et al. Comparative Gene Expression Analyses Identify Luminal and Basal Subtypes of Canine Invasive Urothelial Carcinoma That Mimic Patterns in Human Invasive Bladder Cancer. PLoS One. 2015; 10(9):e0136688 [PubMed] Free Access to Full ArticleRelated Publications
More than 160,000 people are expected to die from invasive urothelial carcinoma (iUC) this year worldwide. Research in relevant animal models is essential to improving iUC management. Naturally-occurring canine iUC closely resembles human iUC in histopathology, metastatic behavior, and treatment response, and could provide a relevant model for human iUC. The molecular characterization of canine iUC, however, has been limited. Work was conducted to compare gene expression array results between tissue samples from iUC and normal bladder in dogs, with comparison to similar expression array data from human iUC and normal bladder in the literature. Considerable similarities between enrichment patterns of genes in canine and human iUC were observed. These included patterns mirroring basal and luminal subtypes initially observed in human breast cancer and more recently noted in human iUC. Canine iUC samples also exhibited enrichment for genes involved in P53 pathways, as has been reported in human iUC. This is particularly relevant as drugs targeting these genes/pathways in other cancers could be repurposed to treat iUC, with dogs providing a model to optimize therapy. As part of the validation of the results and proof of principal for evaluating individualized targeted therapy, the overexpression of EGFR in canine bladder iUC was confirmed. The similarities in gene expression patterns between dogs and humans add considerably to the value of naturally-occurring canine iUC as a relevant and much needed animal model for human iUC. Furthermore, the finding of expression patterns that cross different pathologically-defined cancers could allow studies of dogs with iUC to help optimize cancer management across multiple cancer types. The work is also expected to lead to a better understanding of the biological importance of the gene expression patterns, and the potential application of the cross-species comparisons approach to other cancer types as well.
Kato Y, Ochiai K, Michishita M, et al. Molecular cloning of canine co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein α (SGTA) and investigation of its ability to suppress androgen receptor signalling in androgen-independent prostate cancer. Vet J. 2015; 206(2):143-8 [PubMed] Related Publications
Although the morbidity of canine prostate cancer is low, the majority of cases present with resistance to androgen therapy and poor clinical outcomes. These pathological conditions are similar to the signs of the terminal stage of human androgen-independent prostate cancer. The co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein α (SGTA) is known to be overexpressed in human androgen-independent prostate cancer. However, there is little information about the structure and function of canine SGTA. In this study, canine SGTA was cloned and analysed for its ability to suppress androgen receptor signalling. The full-length open reading frame (ORF) of the canine SGTA gene was amplified by RT-PCR using primers designed from canine-expressed sequence tags that were homologous to human SGTA. The canine SGTA ORF has high homology with the corresponding human (89%) and mouse (81%) sequences. SGTA dimerisation region and tetratricopeptide repeat (TPR) domains are conserved across the three species. The ability of canine SGTA to undergo homodimerisation was demonstrated by a mammalian two-hybrid system and a pull-down assay. The negative impact of canine SGTA on androgen receptor (AR) signalling was demonstrated using a reporter assay in androgen-independent human prostate cancer cell lines. Pathological analysis showed overexpression of SGTA in canine prostate cancer, but not in hyperplasia. A reporter assay in prostate cells demonstrated suppression of AR signalling by canine SGTA. Altogether, these results suggest that canine SGTA may play an important role in the acquisition of androgen independence by canine prostate cancer cells.
Bonnefont-Rebeix C, Fournel-Fleury C, Ponce F, et al. Characterization of a novel canine T-cell line established from a spontaneously occurring aggressive T-cell lymphoma with large granular cell morphology. Immunobiology. 2016; 221(1):12-22 [PubMed] Related Publications
Dogs with lymphoma are established as good model for human non-Hodgkin lymphoma studies. Canine cell lines derived from lymphomas may be valuable tools for testing new therapeutic drugs. In this context, we established a canine T-cell line, PER-VAS, from a primary aggressive T-cell lymphoma with large granular morphology. Flow cytometric analysis revealed a stable immunophenotype: PER-VAS cells were positively labelled for CD5, CD45, MHC II and TLR3, and were negative for CD3, CD4 and CD8 expression. Although unstable along the culture process, IL-17 and MMP12 proteins were detectable as late as at passages 280 and 325i.e. respectively 24 and 29 months post isolation. At passage 325, PER-VAS cells maintained the expression of IL-17, CD3, CD56, IFNγ and TNFα mRNAs as shown by RT-PCR analysis. Stable rearrangement of the TCRγ gene has been evidenced by PCR. PER-VAS cells have a high proliferation index with a doubling time of 16.5h and were tumorigenic in Nude mice. Compared to the canine cell lines already reported, PER-VAS cells display an original expression pattern, close to NKT cells, which makes them valuable tools for in vitro comparative research on lymphomas.
Posch F, Thaler J, Zlabinger GJ, et al. Soluble Vascular Endothelial Growth Factor (sVEGF) and the Risk of Venous Thromboembolism in Patients with Cancer: Results from the Vienna Cancer and Thrombosis Study (CATS). Clin Cancer Res. 2016; 22(1):200-6 [PubMed] Related Publications
PURPOSE: Cancer patients are at an increased risk of venous thromboembolism (VTE). VEGF promotes the growth of highly thrombogenic tumor vessels. Here, we explored the utility of soluble plasma VEGF-A (sVEGF) as a biomarker for the prediction of VTE in patients with cancer. EXPERIMENTAL DESIGN: Eight hundred four patients with newly diagnosed cancer or progression after remission were prospectively followed for 2 years until the occurrence of VTE or death [tumor sites: brain (n = 87), breast (n = 137), lung (n = 120), gastrointestinal (n = 143), pancreas (n = 53), prostate (n = 95), kidney (n = 22), myeloma and lymphoma (n = 99), and others (n = 48)]. Primary endpoint was symptomatic or fatal VTE. sVEGF was measured by immunoassay in baseline plasma. RESULTS: Fifty-five patients developed VTE (6.8%) and 364 patients (45.3%) died. Five-hundred and forty-two (68.3%) participants had sVEGF levels above the detection limit of 0.5 pg/mL. The median sVEGF level (25th-75th percentile) was 8.1 pg/mL (0-17.7). The cumulative 2-year incidence of VTE was 10.2% [95% confidence interval (CI), 6.4-14.9] in patients with sVEGF greater than the 75th percentile of the sVEGF distribution (Q3, cutoff: 17.7 pg/mL), and 5.9% (95% CI, 4.2-7.9) in patients with lower levels (P = 0.03). The corresponding 2-year risk of death was 52.8% (95% CI, 46.0-60.0) and 43.9% (95% CI, 40.0-48.0), respectively (P = 0.02). In univariable time-to-VTE regression, elevated sVEGF was associated with VTE [subhazard ratio (SHR) per 10 pg/mL increase, 1.04; 95% CI,1.00-1.09; P = 0.04)]. The association between sVEGF and risk of VTE prevailed after multivariable adjustment for high-risk tumor sites, age, gender, factor VIII, thrombin generation potential, and soluble P-Selectin (adjusted SHR, 1.04; 95% CI, 1.00-1.09, P = 0.05). CONCLUSIONS: Elevated sVEGF is associated with an increased risk of VTE in patients with cancer.